Tjorvi E. Perry

Functional small-diameter neovessels created using endothelial progenitor cells expanded ex vivo

Arterial conduits are increasingly preferred for surgical bypass because of inherent functional properties conferred by arterial endothelial cells, especially nitric oxide production in response to physiologic stimuli. Here we tested whether endothelial progenitor cells (EPCs) can replace arterial endothelial cells and promote patency in tissue-engineered small-diameter blood vessels (4 mm). We isolated EPCs from peripheral blood of sheep, expanded them ex vivo and then seeded them on decellularized porcine iliac vessels. EPC-seeded grafts remained patent for 130 days as a carotid interposition graft in sheep, whereas non-seeded grafts occluded within 15 days. The EPC-explanted grafts exhibited contractile activity and nitric-oxide–mediated vascular relaxation that were similar to native carotid arteries. These results indicate that EPCs can function similarly to arterial endothelial cells and thereby confer longer vascular-graft survival. Due to their unique properties, EPCs might have other general applications for tissue-engineered structures and in treating vascular diseases.

From Stem Cells to Viable Autologous Semilunar Heart Valve

Background—An estimated 275 000 patients undergo heart valve replacement each year. However, existing solutions for valve replacement are complicated by the morbidity associated with lifelong anticoagulation of mechanical valves and the limited durability of bioprostheses. Recent advances in tissue engineering and our understanding of stem cell biology may provide a lifelong solution to these problems. Methods and Results—Mesenchymal stem cells were isolated from ovine bone marrow and characterized by their morphology and antigen expression through immunocytochemistry, flow cytometry, and capacity to differentiate into multiple cell lineages. A biodegradable scaffold was developed and characterized by its tensile strength and stiffness as a function of time in cell-conditioned medium. Autologous semilunar heart valves were then created in vitro using mesenchymal stem cells and the biodegradable scaffold and were implanted into the pulmonary position of sheep on cardiopulmonary bypass. The valves were evaluated by echocardiography at implantation and after 4 months in vivo. Valves were explanted at 4 and 8 months and examined by histology and immunohistochemistry. Valves displayed a maximum instantaneous gradient of 17.2±1.33 mm Hg, a mean gradient of 9.7±1.3 mm Hg, an effective orifice area of 1.35±0.17 cm2, and trivial or mild regurgitation at implantation. Gradients changed little over 4 months of follow-up. Histology showed disposition of extracellular matrix and distribution of cell phenotypes in the engineered valves reminiscent of that in native pulmonary valves. Conclusions—Stem-cell tissue-engineered heart valves can be created from mesenchymal stem cells in combination with a biodegradable scaffold and function satisfactorily in vivo for periods of >4 months. Furthermore, such valves undergo extensive remodeling in vivo to resemble native heart valves.

Variation in the 4q25 Chromosomal Locus Predicts Atrial Fibrillation after Coronary Artery Bypass Graft Surgery

Background— Atrial fibrillation (AF) is the most common adverse event following coronary artery bypass graft surgery. A recent study identified chromosome 4q25 variants associated with AF in ambulatory populations. However, their role in postoperative AF is unknown. We hypothesized that genetic variants in the 4q25 chromosomal region are independently associated with postoperative AF after coronary artery bypass graft surgery. Methods and Results— Two prospectively collected cohorts of patients undergoing coronary artery bypass graft surgery, with or without concurrent valve surgery, at 3 US centers. From a discovery cohort of 959 patients, clinical and genomic multivariate predictors of postoperative AF were identified by genotyping 45 single-nucleotide polymorphisms (SNPs) encompassing the 4q25 locus. Three SNPs were then assessed in a separately collected validation cohort of 494 patients. After adjustment for clinical predictors of postoperative AF and multiple comparisons, rs2200733, rs13143308, and 5 other linked SNPs independently predicted postoperative AF in the discovery cohort. Additive odds ratios for the 7 associated 4q25 SNPs ranged between 1.57 and 2.17 (P=8.0×10−4 to 3.4×10−6). Association with postoperative AF were measured and replicated for rs2200733 and rs13143308 in the validation cohort. Conclusions— In 2 independently collected cardiac surgery cohorts, noncoding SNPs within the chromosome 4q25 region are independently associated with postoperative AF after coronary artery bypass graft surgery after adjusting for clinical covariates and multiple comparisons.

Dynamic rotational seeding and cell culture system for vascular tube formation.

Optimization of cell seeding and culturing is an important step for the successful tissue engineering of vascular conduits. We evaluated the effectiveness of using a hybridization oven for rotational seeding and culturing of ovine vascular myofibroblasts onto biodegradable polymer scaffolds suitable for replacement of small- and large-diameter blood vessels. Large tubes (12 mm internal diameter and 60 mm length, n = 4) and small tubes (5 mm internal diameter and 20 mm length, n = 4) were made from a combination of polyglycolic acid/poly-4-hydroxybutyrate and coated with collagen solution. Tubes were then placed in culture vessels containing a vascular myofibroblast suspension (10(6) cells/cm(2)) and rotated at 5 rpm in a hybridization oven at 37 degrees C. Light and scanning electron microscopy analyses were performed after 5, 7, and 10 days. Myofibroblasts had formed confluent layers over the outer and inner surfaces of both large and small tubular scaffolds by day 5. Cells had aligned in the direction of flow by day 7. Multiple spindle-shaped cells were observed infiltrating the polymer mesh. Cell density increased between day 5 and day 10. All conduits maintained their tubular shape throughout the experiment. We conclude that dynamic rotational seeding and culturing in a hybridization oven is an easy, effective, and reliable method to deliver and culture vascular myofibroblasts onto tubular polymer scaffolds.

Plasma Neutrophil Gelatinase-Associated Lipocalin and Acute Postoperative Kidney Injury in Adult Cardiac Surgical Patients

BACKGROUND: Acute kidney injury (AKI) after coronary artery bypass graft (CABG) surgery is associated with increased postoperative morbidity and mortality. We hypothesized that increased plasma neutrophil gelatinase-associated lipocalin (NGAL) measured immediately after separating from cardiopulmonary bypass (CPB) would predict AKI after CABG surgery. METHODS: In a retrospective observational study, we examined the value of plasma NGAL measured after CPB for predicting the risk of developing AKI (defined as a ≥50% increase in serum creatinine from preoperative levels) in 879 patients after CABG surgery using multivariable logistic regression. Area under the curve of receiver operating characteristic curves was analyzed to assess sensitivities, specificities, and cutoff points for postoperative plasma NGAL levels to predict AKI. RESULTS: Seventy-five patients (8.6%) developed postoperative AKI. Plasma NGAL levels measured after CPB were higher in patients who subsequently developed AKI than in those who did not (AKI: 268.8 ng/mL [207.5–459.5 ng/mL], median [interquartile range], vs no AKI: 238.4 ng/mL [172.0–319.1 ng/mL]; P < 0.001) and remained higher through postoperative day 4. An optimal serum plasma NGAL cutoff of 353.5 ng/mL at the post-CPB time point had a sensitivity of 38.7%, specificity of 81.5%, and a positive predictive value of 16.3% for predicting AKI. In our multivariate regression model, post-CPB plasma NGAL levels >353.5 ng/mL were independently associated with postoperative AKI (odds ratio, 2.3; 95% confidence interval, 1.5–6.5; P = 0.002). CONCLUSION: An early increase of post-CPB plasma NGAL is associated with AKI in adult patients undergoing CABG surgery, although the sensitivity is low. Therefore, assessing early plasma NGAL alone has limited utility for predicting AKI in this patient population.

Natriuretic peptide system gene variants are associated with ventricular dysfunction after coronary artery bypass grafting.

Background:Ventricular dysfunction (VnD) after primary coronary artery bypass grafting is associated with increased hospital stay and mortality. Natriuretic peptides have compensatory vasodilatory, natriuretic, and paracrine influences on myocardial failure and ischemia. The authors hypothesized that natriuretic peptide system gene variants independently predict risk of VnD after primary coronary artery bypass grafting. Methods:A total of 1,164 patients undergoing primary coronary artery bypass grafting with cardiopulmonary bypass at two institutions were prospectively enrolled. After prospectively defined exclusions, 697 patients of European descent (76 with VnD) were analyzed. VnD was defined as need for at least 2 new inotropes and/or new mechanical ventricular support after coronary artery bypass grafting. A total of 139 haplotype-tagging single nucleotide polymorphisms (SNPs) within 7 genes (NPPA, NPPB, NPPC, NPR1, NPR2, NPR3, CORIN) were genotyped. SNPs univariately associated with VnD were entered into logistic regression models adjusting for clinical covariates predictive of VnD. To control for multiple comparisons, permutation analyses were conducted for all SNP associations. Results:After adjusting for clinical covariates and multiple comparisons within each gene, seven NPPA/NPPB SNPs (rs632793, rs6668352, rs549596, rs198388, rs198389, rs6676300, rs1009592) were associated with decreased risk of postoperative VnD (additive model; odds ratios 0.44–0.55; P = 0.010–0.036) and four NPR3 SNPs (rs700923, rs16890196, rs765199, rs700926) were associated with increased risk of postoperative VnD (recessive model; odds ratios 3.89–4.28; P = 0.007–0.034). Conclusions:Genetic variation within the NPPA/NPPB and NPR3 genes is associated with risk of VnD after primary coronary artery bypass grafting. Knowledge of such genotypic predictors may result in better understanding of the molecular mechanisms underlying postoperative VnD.

Comparison of the utility of preoperative versus postoperative B-type natriuretic peptide for predicting hospital length of stay and mortality after primary coronary artery bypass grafting.

Background:Preoperative B-type natriuretic peptide (BNP) is known to predict adverse outcomes after cardiac surgery. The value of postoperative BNP for predicting adverse outcomes is less well delineated. The authors hypothesized that peak postoperative plasma BNP (measured postoperative days 1–5) predicts hospital length of stay (HLOS) and mortality in patients undergoing primary coronary artery bypass grafting, even after adjusting for preoperative BNP and perioperative clinical risk factors. Methods:This study is a prospective longitudinal study of 1,183 patients undergoing primary coronary artery bypass grafting surgery. Mortality was defined as all-cause death within 5 yr after surgery. Cox proportional hazards analyses were conducted to separately evaluate the associations between peak postoperative BNP and HLOS and mortality. Multivariable adjustments were made for patient demographics, preoperative BNP concentration, and clinical risk factors. BNP measurements were log10 transformed before analysis. Results:One hundred fifteen deaths (9.7%) occurred in the cohort (mean follow-up = 4.3 yr, range = 2.38-5.0 yr). After multivariable adjustment for preoperative BNP and clinical covariates, peak postoperative BNP predicted HLOS (hazard ratio [HR] = 1.28, 95% CI = 1.002-1.64, P = 0.049) but not mortality (HR = 1.62, CI = 0.71-3.68, P = 0.25), whereas preoperative BNP independently predicted HLOS (HR = 1.09, CI = 1.01-1.18, P = 0.03) and approached being an independent predictor of mortality (HR = 1.36, CI = 0.96-1.94, P = 0.08). When preoperative and peak postoperative BNP were separately adjusted for within the clinical multivariable models, each independently predicted HLOS (preoperative BNP HR = 1.13, CI = 1.05-1.21, P = 0.0007; peak postoperative BNP HR = 1.44, CI = 1.15-1.81, P = 0.001) and mortality (preoperative BNP HR = 1.50, CI = 1.09-2.07, P = 0.01; peak postoperative BNP HR = 2.29, CI = 1.11-4.73, P = 0.02). Conclusions:Preoperative BNP may be better than peak postoperative BNP for predicting HLOS and longer term mortality after primary coronary artery bypass grafting surgery.

Postoperative Activity, but Not Preoperative Activity, of Antithrombin Is Associated with Major Adverse Cardiac Events After Coronary Artery Bypass Graft Surgery

BACKGROUND:Low levels of antithrombin (AT) have been independently associated with prolonged intensive care unit stay and an increased incidence of neurologic and thromboembolic events after cardiac surgery. We hypothesized that perioperative AT activity is independently associated with postoperative major adverse cardiac events (MACEs) in patients undergoing coronary artery bypass graft (CABG) surgery. METHODS:We prospectively studied 1403 patients undergoing primary CABG surgery with cardiopulmonary bypass (CPB) (http://clinicaltrials.gov/show/NCT00281164). The primary clinical end point was occurrence of MACE, defined as a composite outcome of any one or more of the following: postoperative death, reoperation for coronary graft occlusion, myocardial infarction, stroke, pulmonary embolism, or cardiac arrest until first hospital discharge. Plasma AT activity was measured before surgery, after post-CPB protamine, and on postoperative days (PODs) 1–5. Multivariate logistic regression modeling was performed to estimate the independent effect of perioperative AT activity upon MACE. RESULTS:MACE occurred in 146 patients (10.4%), consisting of postoperative mortality (n = 12), myocardial infarction (n = 108), stroke (n = 17), pulmonary embolism (n = 8), cardiac arrest (n = 16), or a subsequent postoperative or catheter-based treatment for graft occlusion (n = 6). AT activity at baseline did not differ between patients with (0.91 ± 0.13 IU/mL; n = 146) and without (0.92 ± 0.13 IU/mL; n = 1257) (P = 0.18) MACE. AT activity in both groups was markedly reduced immediately after CPB and recovered to baseline values over the ensuing 5 PODs. Postoperative AT activity was significantly lower in patients with MACE than those without MACE. After adjustment for clinical predictors of MACE, AT activity on PODs 2 and 3 was associated with MACE. CONCLUSIONS:Preoperative AT activity is not associated with MACE after CABG surgery. MACE is independently associated with postoperative AT activity but only at time points occurring predominantly after the MACE.

Heart-Type Fatty Acid Binding Protein Is an Independent Predictor of Death and Ventricular Dysfunction After Coronary Artery Bypass Graft Surgery

BACKGROUND: Heart-type fatty acid binding protein (hFABP) functions as a myocardial fatty acid transporter and is released into the circulation early after myocardial injury. We hypothesized that hFABP is superior to conventional cardiac biomarkers for predicting early perioperative myocardial injury after coronary artery bypass graft (CABG) surgery. METHODS: A prospective cohort study of 1298 patients undergoing primary CABG with cardiopulmonary bypass (CPB) was performed at 2 institutions. Four plasma myocardial injury biomarkers (hFABP; cardiac troponin I [cTnI]; creatine kinase, MB [CK-MB] fraction; and myoglobin) were measured at 7 perioperative time points. The association among perioperative cardiac biomarkers and ventricular dysfunction, hospital length of stay (HLOS), and up to 5-year postoperative mortality (median 3.3 years) was assessed using Cox proportional hazard models. We defined in-hospital ventricular dysfunction as a new requirement for 2 or more inotropes, or new placement of an intraaortic balloon pump, or ventricular assist device either during the intraoperative period after the patient separated from CPB or postoperatively in the intensive care unit. RESULTS: The positive and negative predictive values of mortality for hFABP are 13% (95% confidence interval [CI], 9%–19%) and 95% (95% CI, 94%–96%), respectively, which is higher than for cTnI and CK-MB. After adjusting for clinical predictors, both postoperative day (POD) 1 and peak hFABP levels were independent predictors of ventricular dysfunction (P < 0.0001), HLOS (P < 0.05), and 5-year mortality (P < 0.0001) after CABG surgery. Furthermore, POD1 and peak hFABP levels were significantly superior to other evaluated biomarkers for predicting mortality. In a repeated-measures analysis, hFABP outperformed all other models of fit for HLOS. Patients with POD2 hFABP levels higher than post-CPB hFABP levels had an increased mortality compared with those patients whose POD2 hFABP levels decreased from their post-CPB level (hazard ratio, 10.9; 95% CI, 5.0–23.7; P = 7.2 × 10−10). Mortality in the 120 patients (10%) with a later hFABP peak was 18.3%, compared with 4.7% in those who did not peak later. Alternatively, for cTnI or CK-MB, no difference in mortality was detected. CONCLUSION: Compared with traditional markers of myocardial injury after CABG surgery, hFABP peaks earlier and is a superior independent predictor of postoperative mortality and ventricular dysfunction.

Preoperative C-reactive Protein Predicts Long-term Mortality and Hospital Length of Stay after Primary, Nonemergent Coronary Artery Bypass Grafting

Background:Preoperative C-reactive protein (CRP) levels more than 10 mg/l have been shown to be associated with increased morbidity and mortality after cardiac surgery. We examine the value of preoperative CRP levels less than 10 mg/l for predicting long-term, all-cause mortality and hospital length of stay in surgical patients undergoing primary, nonemergent coronary artery bypass graft-only surgery. Methods:We examined the association between preoperative CRP levels stratified into four categories (< 1, 1-3, 3-10, and > 10 mg/l), and 7-yr all-cause mortality and hospital length of stay in 914 prospectively enrolled primary, nonemergent coronary artery bypass graft-only surgical patients using a proportional hazards regression model. Results:Eighty-seven patients (9.5%) died during a mean follow-up period of 4.8 ± 1.5 yr. After proportional hazards adjustment, the 3-10 and > 10 mg/l preoperative CRP groups were associated with long-term, all-cause mortality (hazards ratios [95% CI]: 2.50 [1.22-5.16], P = 0.01 and 2.66 [1.21-5.80], P = 0.02, respectively) and extended hospital length of stay (1.32 [1.07-1.63], P < 0.001 and 1.27 [1.02-1.62], P = 0.001, respectively). Conclusion:We demonstrate that preoperative CRP levels as low as 3 mg/l are associated with increased long-term mortality and extended hospital length of stay in relatively lower-acuity patients undergoing primary, nonemergent coronary artery bypass graft-only surgery. These important findings may allow for more objective risk stratification of patients who present for uncomplicated surgical coronary revascularization.